Maturation of the Coxiella burnetii parasitophorous vacuole requires bacterial protein synthesis but not replication

Abstract

SummaryThis study examined whether protein synthesis and replication are required for maturation and fusogenicity of the lysosomal-like, large and spacious parasitophorous vacuole (PV) of Coxiella burnetii , an obligate intracellular bacterium. Large and spacious PV with multiple non-replicating C. burnetii were observed by phase microscopy in Vero cells infected at a multiplicity of infection of ten and treated with a bacteriostatic concentration of nalidixic acid or carbenicillin, antimicrobics that inhibit DNA and cell wall biosynthesis respectively. Conversely, large and spacious PV were not observed in cells treated with a bacteriostatic concentration of the protein synthesis inhibitor chloramphenicol. Rather, fluorescence microscopy of individual cells revealed multiple, acidic PV harbouring a single organism tightly bounded by a LAMP-1 positive vacuolar membrane. These vacuoles homotypically fused to form a large and spacious PV upon removal of the drug. Chloramphenicol also inhibited trafficking of latex beads to large and spacious PV and caused mature PV to collapse. Collectively, these results demonstrate that C. burnetii protein synthesis, but not replication, is required for fusion between nascent C. burnetii PV and latex bead phagosomes, and also for formation and maintenance of large and spacious, replicative PV. However, transit of nascent PV through the endocytic pathway to ultimately acquire lysosomal markers appears to occur irrespective of Coxiella protein synthesis.

VirulentCoxiella burnetiidoes not activate human dendritic cells: Role of lipopolysaccharide as a shielding molecule

“…Fig. 4A shows that both strains formed large, spacious, CD63-positive vacuoles that appear indistinguishable from vacuoles formed in other cell types known to be permissive for C. burnetii growth (31,32).…”

Coxiella burnetii effector protein subverts clathrin-mediated vesicular trafficking for pathogen vacuole biogenesis

“…However, C. burnetii modulation of PV fusogenicity is also considered essential for successful infection. Cell culture infection models have revealed several host vesicular pathways involved in PV biogenesis (5,7,25). Disruption of Rab GTPases that regulate endocytic (Rab5 and Rab7), secretory (Rab1), and autophagic (Rab24) vesicular trafficking events produce defects in intracellular replication (5,8,26,27), implying that C. burnetii obtains lipids and proteins for PV biogenesis, as well as nutrients for growth, from heterotypic fusion with multiple vesicular compartments.…”

“…Like other intracellular bacteria, C. burnetii actively modifies its intracellular niche, or parasitophorous vacuole (PV). Bacterial protein synthesis is required for homotypic and heterotypic fusion of the PV with cellular vesicles to result in a replication compartment that can occupy nearly the entire host-cell cytoplasm (5)(6)(7)(8). However, the C. burnetii PV is unique among bacteria-occupied vacuoles by resembling, in structure and function, a large phagolysosome (2).…”

Is Q Fever an emerging or re-emerging zoonosis?

“…Moreover, the small forms are able to delay phagolysosomal fusion, perhaps to facilitate the transition from SCV to LCV that occurs at pH 5.5 in the endosome [66]. The use of cellular markers (LAMP-1, EEA.1, rab7) [67,68] strongly suggests that the large replicative vacuoles containing Coxiella are derived from fusion with late endosomal-lysosomal organelles. Sequence homologies between the type IV secretion system proteins of Coxiella and Legionella strongly suggest that this secretion system should be involved in the maturation of the phagosome containing Coxiella [144].…”